This is a proposal to investigate the role of calcium in the morphological and functional differentiation of skeletal muscle. For this purpose, electrophysiological, ultrastructural and microscopic fluorometric methods will be applied to cultured muscle cells (myotubes) from normal mice and from mice with the muscular dysgenesis mutation. This mutation alters the structural gene for the skeletal muscle dihydropyridine (DHP) receptor, a protein that appears to function both as a slow calcium channel and as an essential component of excitation-contraction (E-C) coupling. Due to the absence of the DDP receptor, dysgenic muscle is paralyzed and fails to develop normal structure. "Chimeric", dysgenic myotubes will be used as model system to examine the function of calcium in muscle differentiation. These chimeric myotubes, which contain a single normal nucleus, recover E-C coupling and normal structure, but only in the region of the normal nucleus. Cytosolic calcium transients will be quantified, before and after nucleus. The timing and spatial extent of ultrastructural recovery in elevated calcium. To determine the effective lifetime of slow calcium after destruction of the normal nucleus in a chimeric myotube. To further test the role of cytosolic calcium in producing sarcomeric several alternative methods, including the expression of exogenous calcium channels by injection of cDNAs into dysgenic myotubes. The spatial localization of different kinds of calcium channels will be compared. The proposed experiments will provide new insights into the way in which calcium channel subtypes are regionally distributed and into how these channels and cytosolic calcium regulate cellular differentiation.